This study took place in a malaria-endemic region of Mali in West Africa at Hospital Gabriel Tour in Bamako, which is the major pediatric referral hospital in Mali. The first subject was enrolled on 10/20/08. As of 6/25/08, 102 participants (50 males, 52 females) in Mali had been enrolled into the study. The mean age was 32 months. A standardized history and physical examination was performed. Each participant also underwent echocardiography (165 completed echocardiograms including follow-ups). Obtaining a blood sample for standard hematology and chemistry panels, research bloods, as well as DNA collection for isolation and plasma storage followed the echocardiogram. Based on these studies the prevalence of PAH in the severe malarial anemia subjects was 64% versus 19% in the healthy controls. Children with severe malaria had higher plasma levels of hemoglobin and arginase-1, reduced wholeblood levels of nitrite, and increased NO consumption relative to controls. They also had increased pulmonary arterial pressures (P<.05) with elevated levels of NT-proBNP and soluble vascular cell adhesion molecule-1 (P<.001). Children with severe malaria have increased pulmonary pressures and myocardial wall stress. These complications are consistent with NO depletion from intravascular hemolysis, and they indicate that the pathophysiologic cascade from intravascular hemolysis to NO depletion and its cardiopulmonary effects is activated in children with severe malaria. We also have echocardiographs on the asymptomatic parasitemia and uncomplicated malaria arms, but this data has not yet been analyzed. We have performed HPLC analysis of 181 plasma samples from 97 subjects. In summary, we have found arginine depletion and relative elevation of methylarginines in plasma of children with acute malaria infection. The shift in arginine bioavailability would limit NO synthesis and increased generation of reactive oxygen species. This confirms our prior observations of arginine depletion in Gambian children with malaria. Arginine depletion is though to be caused by increased arginase activity that converts arginine to ornithine. If so, then ornithine levels should be elevated. However, we found surprisingly that ornithine also decreased with malaria in infection. Further analysis of citrulline levels confirmed that malaria infection caused all three major urea cycle intermediates arginine, ornithine and citrulline to be diminished. This implies that the carbon skeletons from these molecules are being directed towards other metabolic fates such as energy production or polyamine synthesis, rather then simply increased conversion from arginine to ornithine. To pursue the novel hypothesis that arginine depletion is caused by increased flux of amino acids out of the urea cycle to alternative metabolic fates, we conducted quantitative studies of arginine metabolism using heavy isotope labeled amino acids in a mouse model of severe malaria. The goal of these experiments was to quantify the changes in amino acid metabolism that occur during malaria infection in a controlled model system. First we determined that genetic knock-out of parasite arginase did not prevent arginine depletion in the infected mouse. Next we confirmed that conversion of arginine to ornithine by arginase was unchanged by malaria infection. This implies that arginine depletion during a malaria infection is not due to high arginase activity, as previously proposed. Instead it is a decreased rate of appearance of arginine in plasma that is responsible for low plasma arginine levels in malaria-infected mice. This implies that arginine synthesis release of arginine from cells is diminished during an acute malaria infection. This suggests that arginine supplementation may be successful at raising arginine levels and improving NO synthesis in the host. All remaining de-identified samples and data from this study were transferred via a Material Transfer Agreement to The Translational Research Core Laboratory at the Division of Pulmonary, Allergy and Critical Care Medicine at the University of Pittsburgh Medical Center to be housed in the Biological specimen repository for the Hemolysis, NO and PAH in malaria study. This transfer took place in May 2015.